In the nonobese diabetic (NOD) mouse, insulin dependent diabetes is an autoimmune disease characterized by T cell-mediated invasion and destruction of pancreatic islet beta cells. The importance of insulin receptor (IR) expression in the pathogenesis of diabetes was examined, since it has been shown that the IR is a chemotactic receptor capable of directing cell movement in response to insulin. Using polyclonal antisera to the IR, phenotypic analysis of purified splenic T cells from diabetic mice showed that on average 15% of T cells expressed high density IR (IR HI). In addition, IR HI T cells were already a dominant phenotype in the insulitis of young prediabetic mice. To determine the ability of IR HI T cells to transfer diabetes, cells were sorted by flow cytometry before adoptive transfer into prediabetic irradiated NOD mice. Transfer of as few as 3x106 purified IRHI T cells alone resulted in rapid onset insulitis and diabetes, whereas IRHI depleted T cells were essentially unable to passage either insulitis or diabetes. Disease was not due to the transfer of activated cells, since removal of IL-2R+ or transferrin R+ cells did not alter diabetes transfer. Therefore, IRHI T cells are aggressively diabetogenic, suggesting that increase IR expression may provide a mechanism for delivering potentially autoreactive T cells to the islet, regardless of their activation state. This grant seeks to further characterize IRHI T cells and determine if IR expression on the surface of T cells in important for diabetogenicity. Initially monoclonal antibodies to the mouse IR will be isolated to provide an unlimited sources of antibodies for these studies. IRHI T cells will be characterized by: 1) determining the ability of IRHI T cells to migrate in response to insulin or islet cells in chemotaxis assays and 2) determining the reactivity of IR HI T cells for insulin and islet cells by proliferation and cytotoxicity assays. The importance of IR expression for pathogenicity will be analyzed by three methods. One, determining the levels of IRHI T cells in the islet by immunohistochemical staining, as NOD mice progress to diabetes. Two, determining if expression of IR or modulation of IR expression on known diabetogenic T cell clones has an effect on the ability to transfer disease. Three, determining if IRHI T cells in the spleens or islets of prediabetic mice are capable of passaging diabetes compared to IRLO T cells from the same sources. This would pinpoint early pathogenic cells. Normal and autoimmune strains of the mice will also be analyzed for IRHI T cells. If present, islet and insulin reactivity will be tested since IR expression may facilitate movement into the pancreas., but antigenic specificity would be required for beta cell destruction. IR expression on the surface of T cells is inversely related to insulin levels in the blood. This could explain why pretreatment of NOD mice with insulin prevents diabetes. If the IRHI T cells are pathogenic, then down regulation of these receptors may prevent movement toward the islet and thus beta cell destruction. This is supported by our preliminary data showing that IRLO T cells, obtained from diabetic mice, do not cause insulitis or diabetes. These experiments will generate novel information on diabetogenic T cells and may elucidate mechanisms for control.